The Cellular UPS, Dr. Octopus, & a Hand-Off

Cells are often described as factories, and their product is protein. Thousands of different proteins are built by cellular structures called ribosomes, which translate DNA instructions into chains of amino acids. But in a cell, as in industry, manufacturing is only the first part of the story: products must also be shipped to their final destination. Within the cell, that’s often the membrane, the site where many proteins are deposited to perform their functions. Studying these delivery systems – the postal service of the cell – is an important pursuit of cell biologists.

In the last few years, scientists have discovered that there are actually two separate routes to deliver eukaryotic proteins from the ribosome to the endoplasmic reticulum membrane. For most of these proteins, a system discovered in the 1970s known as the co-translational system does the job. But for a certain type of membrane protein, called ‘tail-anchored’ proteins, a specialized delivery pathway exists – call it the UPS to the rest of the cell’s postal service.

Tail-anchored proteins make up only 5 percent of the total inventory of membrane proteins, but even that small slice represents hundreds of biologically important products, Keenan said. If you genetically delete one of the components in the trafficking of these proteins to the membrane in mice, it has catastrophic consequences, killing the animal before it is even born.

“These things play all sort of important roles in a variety of different cellular functions,” Keenan said. “If you screw this pathway up, bad things will happen. At that level they are just fundamentally important.”

Previous studies from the Keenan/Hegde collaboration and other laboratories had identified the key components of a tail-anchored protein transport pathway. In yeast, these include a soluble protein called Get3 and two membrane-bound ‘receptor’ proteins, called Get1 and Get2. But until the current paper, nobody had tested whether these three pieces alone were sufficient to ship a protein from ribosome to membrane. To try this, the team (led by postdoctoral researchers Malaiyalam Mariappan and Agnieszka Mateja) created an artificial system that only contained the three Get proteins and a tail-anchored protein for cargo. To their delight, this streamlined system worked, targeting and inserting the proteins in their proper position.

“We have a minimal system, completely purified, that’s only three components plus the substrate,” Keenan said. “Now we can basically do whatever we want. We can make mutants or chemical modifications, and then we can reconstitute the system and ask, does it work? And if it doesn’t work, we can ask where in this process does it actually fail, and why.”

The researchers also obtained structural information about the dynamic shape of the Get proteins, creating the highest resolution model to date of how a tail-anchored protein is shipped to the membrane. A simple guide to the journey, as it now stands:

1. A complex of two Get3’s bound to two molecules of ATP form a “groove” of the right size and chemical properties to capture a tail-anchored protein (the “substrate”) in the cytosol.

2. Once the substrate is safely nestled in the groove, “hooks” on the end of Get2 grab the complex, and bring it to the membrane. The long, flexible arms of Get2 allow it to function in a way that Keenan jokingly says is “like Dr. Octopus.”

3. Next, Get2 executes a football-style hand off to the adjacent Get1 protein. Binding to Get1 causes the two Get3s to partially “unzip,” wedging open the groove and releasing the tail-anchored substrate for insertion into the membrane.

4. Finally, new ATP molecules bind to Get3 causing it to zip back up into the closed form. This releases it from Get1 so that it can initiate another round of protein delivery in the cytosol.

There is still much to be learned and tested about this model, especially about how the released tail-anchored protein finally is inserted into the membrane. But now that the broad outline has been sketched out and the experimental tools are in place, further work can fill in the details. Keenan also said that studying the process of tail-anchored protein delivery can help inform studies of the co-translational system, which carries many similarities despite being much more complex, with many working parts instead of just three. Farther down the road, learning how to manipulate these systems could help fight disease, including those caused by certain viruses (which may hijack the system for their own nefarious ends), or be helpful in bioengineering.

“The more we understand about different targeting pathways, the better our ability to successfully target proteins where we want,” Keenan said. “Right now, there’s no killer app, but you can imagine a lot of potential uses.”